Result

“In mouse striatal progenitors a target band was detected by STJ97460 PPAR-ϒ Antibody. Double band was detected at ~75 kDa. The molecular weight of PPAR-ϒ (isoforms PPAR-ϒ-1 and PPAR-ϒ-2) is 50-55 kDa .” - K. Palm, Protobios.

Gel electrophoresis information：10% polyacrylamide gel, electric current 35 mA, 90 minutes.

Transfer information：0.45 um PVDF membrane, 1 piece of gel (wet transfer), electric current 350 mA 95 minutes.

Lane No. Antigen Loading amount

Primary antibody

Primary antibody

dilution ratio

Secondary

antibody dilution ratio

Target band KD Visualizati

on time

1 Striatal

progenitors 10 ug

STJ95200 Anti-PPAR-α

antibody 1:1000 1:10000 ~50KD 1 minutes

2

Sriatal progenitors

treated with 20 nM troglitazone

10 ug STJ95200 Anti-

PPAR-α antibody

1:1000 1:10000 ~50KD 1 minutes

Cell Line: Mouse striatal progenitors

Method of validation: Western Blot

Primary Antibody: STJ97460 Anti-PPAR-γ

antibody

Secondary Antibody HRP goat anti-rabbit

Reference antibody: LaminB1 (Abcam)

Dilution ratio: 1:1000

Protocol

Treatment of materials:

Cells were cultured to 70%-80% plating density, nuclear fraction was extracted by standard RIPA buffer with protease inhibitors (afore cytoplasmic fraction was extracted by hypotonic buffer), 5X loading buffer was added, denaturation was performed at 95 C̊ for 5 minutes.

Gel electrophoresis: 10% polyacrylamide gel, electric current 35 mA, 90 minutes.

Transfer: 0.45 um PVDF membrane, 1 piece of gel (wet transfer), electric current 350 mA 90 minutes.

Blocking: 1X TBS Tween (0,1%) with 5% skimmed milk powder for 1 hour.

Primary antibody probing: primary antibody was added to the solution of 1X TBS Tween (0,1%) with 5% skimmed milk powder, 4C̊ for overnight.

Membrane wash: 1X TBS Tween (0.1%) wash for 3 times.

Secondary antibody probing: secondary antibody was added to the solution of 1X TBS Tween (0,1%) with 5% skimmed milk powder with 1:10000 dilution ratio, for 1 hour.

Membrane wash: 1X TBS Tween (0.1%) wash for 3 times.

Visualization: ECL visualization for 1 minute with SuperSignal West Femto ECL-kit (Thermo scientific)

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Figure: line 1 control, line 2 treatment of 20 nM

troglitazone